The formation of hydrogen peroxide and free radicals from (glutathione) GSH is dependent on the activity of g-glutamyltranspeptidase (GGT), an enzyme that is frequently present in high amounts in preneoplastic cells. GSH, in the presence of GGT, can induce lipid peroxidation in vitro, using linolenic acid or linoleic acid as substrates, and in situ in cultured human hepatoma cells. The reaction requires iron, a chelator, GSH, and GGT, is accelerated by GGT activation factors, and is inhibited by specific GGT inhibitors and free-radical scavengers. Peroxidation occurs in the presence of physiological concentrations of iron, and endogenous chelators (citrate, ADP, transferrin). Unlike the mutagenic effect of GSH which is mediated through the formation of H202, lipid peroxidation appears to be effected through a free radical reaction, without the intermediate formation of H202. The effects of antioxidants and free radical scavengers on mutagenicity and lipid peroxidation induced by other sulfhydryls, such as D- and L-cysteine and D- and L-penicillamine will be studied. Studies are in progress to test whether lipid peroxidation can be induced in GGT- rich preneoplastic foci in rodents when challenged with GSH. Studies are under way to validate the mutagenicity results of GSH in AS52 cells, optimize the test protocol, and to the examine the effects of antioxidants and free radical scavengers on GSH mutagenicity. These cells will be used to test the mutagenicity of other substances known, or hypothesized, to form oxygen radicals.